tbuckley/cabot-search huggingface dataset:
Abstract P3038: Novel MicroRNAs Targeting Myosin Binding Protein-C3 Gene In Hypertrophic Cardiomyopathy
Authors: Kalyani Ananthamohan, Akhil Baby, Anil Jegga, Sakthivel Sadayappan
Journal: Circulation Research (2023)

Background: Mutations in cardiac myosin binding protein-C3 ( MYBPC3 ) constitute a significant cause of hypertrophic cardiomyopathy (HCM) with incomplete penetrance, and later age of onset. MYBPC3 variant with 25 base pair deletion in intron 32 ( MYBPC3 Δ25bp ) leads to exon 33 skipping and 3’-untranslated region truncation. Independent transcriptomics studies in HCM individuals carrying c.927-2A&gt;G, c.2373insG, c.3624delC and c.3369_3370insC MYBPC3 variants reported altered levels of cardiac and circulating microRNAs (miRNAs). However, the miRNA-mediated post-transcriptional regulation of the MYBPC3 and MYBPC3 Δ25bp in the HCM phenotype remains largely unknown. Methods: Using a computational consensus approach, top-scoring cardiac-specific miRNAs predicted to target wild-type MYBPC3 (WT) and MYBPC3 Δ25bp were identified. To further characterize the potential regulators, the miRNAs with earlier reports on cardiac and skeletal diseases were selected and their targets were intersected with a compiled set of HCM and heart failure (HF) candidate genes. The overlapping genes were subjected to enrichment analysis for biological processes. Results: The computational analyses predicted 7 high scoring miRNAs for MYBPC3, among which miR-146b-3p, miR-2355-5p, miR-4508, miR-4800-5p and miR-214-5p were predicted to regulate both the WT and MYBPC3 Δ25bp variant, whereas the binding sites for miR-5588-5p, and miR-6500-3p were lost in MYBPC3 Δ25bp variant. In specific, miR-146-3p, miR-214-5p, and miR-2355-5p have been reported to be associated with skeletal muscle dysfunction, HCM and myocardial ischemia-reperfusion injury, respectively. The putative targets of these 3 miRNAs intersected with HCM and HF candidate genes, in addition the intersected targets displayed enrichment for cardiac disease-associated biological processes. Conclusions: Computational prediction suggests that predicted MYBPC3 miRNAs affect biological processes and pathways associated with HCM and HF. Importantly, MYBPC3 Δ25bp show a loss of miRNA regulation compared to the WT. Further studies and experimental validations are required to determine the influence of loss of miRNA regulation in the MYBPC3 Δ25bp and their impact on HCM interventions.
Abstract P2063: Cellular Fusion Drives Polyploidization In Human Cardiomyocytes
Authors: Indu Sivankutty, Lucy Jung, August Y Huang, Sarah Araten, Nazia Hilal, Christopher Walsh, Eunjung Alice Lee, Ming Hui Chen, Sangita Choudhury
Journal: Circulation Research (2023)

Polyploidization is the roadblock to cardiac regeneration. Although endoreplication has been considered to be the main source of polyploidization in cardiomyocytes, recent evidence indicates transient cardiomyocyte fusion and its potential role in cardiac development in zebrafish. To understand polyploidization, it is essential to trace the lineage of cardiomyocytes. The existing lineage method is not suitable for studying the lineage in human heart cardiomyocytes. However, the accumulation of clonal somatic DNA mutation can be traced, and the clonal structure of these somatic mutations can be used for lineage reconstruction of human cardiomyocytes as these non-inherited clonal somatic mutations record the unique history of each somatic cell originating from a zygote and can be used as “endogenous barcodes.” Our analysis of ultra-deep targeted sequencing of ≈500 single cardiomyocyte nuclei using a panel of 253 validated clonal sSNVs revealed that 75.7% of the sequenced cardiomyocytes could be classified into one of the nine clades categorized based on the presence or absence of these mutations. Interestingly, we found that cardiomyocytes can have mutations from more than one clade, indicating the possibilities of cellular fusion in cardiomyocytes. Our results suggest that at least ~10% of tetraploid cardiomyocytes are generated from the fusion mechanism during development, whereas 60% higher ploidy cardiomyocytes are generated from the fusion. Further, we performed ATAC Seq and transcriptome analysis to find out the types of cells that are fused to generate polyploid cardiomyocytes in the human heart. Analysis of the transcriptome and chromatin peaks of tetraploid and higher ploidy cells unraveled a population of cells (cluster 7) that expressed chromatin peaks for endothelial markers (EGFL7) as well as cardiomyocyte-specific markers like TBX5, PLN, etc. Our study demonstrates cellular fusion in the adult human heart and the cell types involved in this fusion event. Understanding the generation of polyploid cardiomyocytes in the human heart might enable us to advance the mechanistic understanding of cardiomyocyte regeneration with the long-term goal of stimulating therapeutic cardiac regeneration.
Abstract P2183: Validation Of A Methodology For Left Ventricular Global Longitudinal Strain Measured With Deformation Encoding Mri To Detect Cardiotoxicity
Authors: Julia Kar, Michael V Cohen, Cherie Revere, Angela Judice, Samuel McQuiston, Christopher Malozzi
Journal: Circulation Research (2023)

This study sought to validate left-ventricular (LV) global longitudinal strain (GLS) derived from an MRI deformation encoding technique for detecting cancer therapy-related cardiac dysfunction (CTRCD) in breast cancer patients. Displacement Encoding with Stimulated Echoes (DENSE) cardiac MRI and 2D echocardiographic images for validation were acquired on 32 breast cancer patients at baseline and 3- and 6-month follow-ups after chemotherapy with antineoplastic agents. The DENSE images were processed with automated Artificial Intelligence segmentation techniques for chamber quantification and 3D strain analysis. The corresponding echocardiographic data were processed with 2D speckle-tracking echocardiography (2D-STE). To establish the reliability of the DENSE-AI tool, intraclass correlation coefficients (ICC) were computed between DENSE and 2DE-derived GLS and other parameters at each time point with Cronbach’s Alpha (C-α) indices. CTRCD risk analyses were conducted on the DENSE-AI and 2D-STE measured parameters with Cox proportional hazard (PH) regression models. The ICC reliability results between DENSE and 2DE-derived GLS were C-α = 0.89 at baseline, C-α = 0.93 at 3 months, and C-α = 0.94 at 6 months. Both DENSE and 2D-STE GLS worsened from baseline to the 3- and 6-month follow-ups (-19.1 ± 2.1% vs. -16.0 ± 3.1% vs. -16.1 ± 2.9%, P&lt;0.001) (Fig. 1). Cox regression showed the 3-month DENSE (HR-per-SD: 2.1; 95% CI: 1.4-3.1, P&lt;0.001) GLS measurements as an independent prognosticator of CTRCD occurrence. In conclusion, LV GLS measurement with the DENSE-AI tool was validated to detect CTRCD in breast cancer patients treated with antineoplastic chemotherapy.
Abstract P2175: Induced Pluripotent Stem Cell-derived Cardiac Myocytes To Assess Anthracycline Cardiotoxicity
Authors: Johanna E Jacobs, Robin Duelen, Hilde Gillijns, Angad Johar, Ward Heggermont, Maurilio Sampaolesi, Heidi Segers, Joerg Herrmann, Lucas Van Aelst, Stefan P Janssens
Journal: Circulation Research (2023)

Background: Anthracyclines are a class of highly effective chemotherapeutics, yet their use is regularly complicated by anthracycline-induced cardiotoxicity (AIC). The advent of induced pluripotent stem cell-derived cardiac myocytes (iPSC-CMs) offers the potential for disease modeling and prediction of genetic cardiomyopathies. Purpose: We aim to use iPSC-CMs to assess susceptibility to AIC. Methods: Five AIC heart transplant patients and five anthracycline treated patients without AIC were included in our study. We reprogrammed lymphocytes from both groups toward iPSCs using Sendai Virus, followed by differentiation into CM-like cells. Cytotoxicity following chemotherapy exposure was assessed using flow cytometry (FC), electron microscopy (EM) and reactive oxygen species generation (ROS). Furthermore, whole genome sequencing (WGS) was performed and variants were filtered based on allele frequency, pathogenicity using HumVar trained variant effect predictors and presence in patients versus controls. Results: The cytotoxic effect of increasing concentrations of doxorubicin was quantifiable using FC on SIRPA positive iPSC-CMs ( A ). There was a higher rate of cell death at any concentration of doxorubicin in patients with AIC as opposed to controls ( B ). On the ultrastructural level, doxorubicin treated iPSC-CMs manifested darker and more condensed mitochondria as seen with EM ( C ). Increased ROS production was observed following doxorubicin exposure ( D ). WGS analysis showed 2 pathogenic variants in the PRAMEF6 gene present in cases and not in controls ( E ). Conclusion: iPSC-CMs represent a robust disease model to study mechanisms of and susceptibility to AIC.
Abstract P1080: Periostin Is Induced During Cell-based Repair In The Sheep Model Of Myocardial Infarction
Authors: Richard Vega, Lorelei Hughes, Nahidh Hasaniya, Leonard Bailey, Mary K Kearns-Jonker
Journal: Circulation Research (2023)

Cardiac disease is the leading cause of death worldwide. Stem cell-based therapy may improve patient outcomes following a myocardial infarction. Our laboratory is studying the mechanistic basis by which neonatal Islet-1 positive cardiovascular progenitor cells (CPC) facilitate post infarct repair in the sheep model of myocardial infarction. Here, we addressed the hypothesis that periostin (POSTN) is induced in the cardiovascular repair zone following neonatal Islet-1+ cell-based repair. POSTN promotes cardiomyocyte proliferation, recruitment of M2 macrophages, epithelial-mesenchymal transition (EMT), migration, and angiogenesis. We examined human neonatal Islet-1+ cardiovascular progenitor cell clones isolated from several patients to initially determine whether or not POSTN was expressed in these cells. All clones expressed POSTN as shown by RT-PCR. The size of the amplified transcripts was confirmed by gel electrophoresis and POSTN expression in these clones was further confirmed by RNA sequencing. We then examined POSTN expression in the cardiovascular repair zone following myocardial infarction and cell transplantation in sheep. Sheep neonatal Islet-1+ CPC clones were used in this model of allogeneic cell-based repair. A myocardial infarction was induced in three Western sheep by left anterior descending coronary artery ligation. Ten million neonatal sheep Islet1+ cardiac progenitor cell clones isolated from 9 day old sheep were injected into the area surrounding the infarct site 3-4 weeks post infarction. RNA was isolated from the cardiovascular repair zone two months after cell injection. The non-infarcted region of the left ventricle and the left ventricle of normal, non-infarcted sheep were used as controls. POSTN transcripts were significantly elevated in the cell-treated cardiovascular repair zone of all three sheep, as identified by qRT-PCR relative to controls. Periostin levels were elevated between 6 and 255 fold in the repair zone relative to the non-infarcted region of the heart. Our data shows that POSTN is expressed in human Islet-1+ neonatal cardiovascular progenitor cells and that POSTN levels are induced during cell-based cardiovascular repair in the sheep model of myocardial infarction.
Abstract EC.07: Tbx5 Maintains Atrial Identity By Regulating An Atrial Enhancer Network
Authors: Mason E Sweat, Yangpo Cao, Xiaoran Zhang, Ozanna Burnicka-Turek, Carlos Perez-Cervantes, Brynn Akerberg, Qing Ma, Hiroko Wakimoto, Josh Gorham, Mi-Kyoung Song, Michael Trembley, Peizhe Wang, Fujian Lu, MATTEO GIANESELLI, Maksymilian K Prondzynski, Raul Bortolin, Jonathan G Seidman, Christine E Seidman, Ivan Moskowitz, William T Pu
Journal: Circulation Research (2023)

Atrial cardiomyocytes (aCMs) and ventricular cardiomyocytes (vCMs) are distinct cell types, but the mechanisms underlying these differences are not well understood. By selectively inactivating the transcription factor Tbx5 in the atrial working myocardium of the neonatal mouse heart, we show that it is required to maintain atrial identity. aCM inactivation of Tbx5 led to the downregulation of highly chamber specific genes including Myl7 and Nppa , and conversely, increased the expression of ventricular identity genes including Myl2 . Using concurrent single nucleus RNA- and ATAC- sequencing, we examined genomic accessibility changes underlying the atrial identity expression program, identifying 1846 genomic loci with greater accessibility in control vs KO aCMs. Nearly 70% of the control-enriched ATAC regions were bound by TBX5, demonstrating a novel role for TBX5 in maintaining atrial genomic accessibility. These regions were associated with genes that had higher expression in control aCMs compared to KO aCMs, suggesting they act as TBX5-dependent enhancers. This hypothesis was tested using HiChIP to analyze enhancer chromatin looping and we determined that 510 chromatin loops were sensitive to TBX5 dosage. Of the loops enriched in control aCMs, 73.7% were anchored in control-enriched ATAC regions. Together, these data demonstrate a role for TBX5 in maintaining atrial identity by supporting chamber specific chromatin confirmation required for aCM specificity.
Abstract P3021: Circulating Exosomal MicroRNAs As Biomarkers For Pulmonary Embolism
Authors: Chongyu Zhang, Srdjan Nikolovski, Bulent Kantarcioglu, Fakiha Siddiqui, Jawed Fareed, Keith Jones
Journal: Circulation Research (2023)

Pulmonary Embolism (PE) is the third leading cause of death worldwide. Current treatment for PE, including thrombolytics and anticoagulants, have serious side effects. Protein and RNA biomarkers presently used to diagnose PE do not have the necessary specificity or sensitivity. Therefore, discovery of predictive biomarkers for therapeutic effects and adverse outcomes is imperative. MicroRNAs (miRNAs) are known to regulate gene expression networks and their expression levels reflect the physiological state of their cell of origin. By investigating these circulating miRNAs, we can gain valuable insight into the state of the damaged cells or tissue. Here, we innovated a novel methodology to isolate exosomes from patient platelet-free plasma that provides better yield of both exosomes and their RNA content. We are the first to stratify the PE patient samples according to clinical severity (low-risk, sub-massive, massive PE) and analyze the correlations of the exosomal miRNA biomarkers with PE severity. We found that exosomes from PE patients are more heterogeneous, lower in concentration and larger in size compared to healthy controls. Further, exosomes from PE patient samples pack more RNA than healthy controls. With the results from ongoing miRNA sequencing, we will be able to identify miRNAs and other short non-coding RNAs as potential diagnostic biomarkers that best correlate with PE severity. Validation and predictive studies using qRT-PCR will push the frontiers on circulating small RNA biomarkers in PE and other cardiovascular diseases. Finally, network analysis on the potential RNA biomarkers will provide information for future studies on disease mechanisms and therapeutics development.
Abstract P2050: Mixed Lineage Kinase 3 Kinase Function Opposes Left Ventricular Hypertrophy And Transcriptional Activity Of Nuclear Factor Of Activated T Cells In The Cardiac Myocyte.
Authors: Loveth Raymond, Gregory Martin, Robert M Blanton
Journal: Circulation Research (2023)

Background: LV hypertrophy (LVH), dysfunction, and remodeling contribute to heart failure (HF). The signaling molecule Mixed Lineage Kinase 3 (MLK3) opposes stress-induced LV dysfunction and remodeling in a kinase-dependent, blood pressure-independent fashion. However, the required downstream MLK3 kinase effectors in the cardiac myocyte (CM) which oppose LVH and remodeling remain poorly understood. The Nuclear Factor of Activated T Cells (NFAT) promotes pathological remodeling, but its regulation in the CM by MLK3 has not been investigated. We hypothesized that MLK3 opposes NFAT in the CM through kinase-dependent mechanisms. Method: In reporter mice with a CM-specific luciferase transgene under the control of NFAT response elements (αMHC-NFAT-Luc mice), we administered the MLK3 inhibitor URMC-099 (10 mg/kg twice daily) or vehicle for 5 days (n=7 vehicle, 8 URMC-099) and 14 days (n=9 vehicle, 12 URMC-099). We measured cardiac chamber masses, performed immunoblotting for MLK3, and measured myocardial luciferase fluorescence as a readout for CM NFAT transcriptional activity. Result: At 5 days, URMC-099 had no effect on LVH or cardiac hypertrophy and did not induce NFAT activation. However, URMC-099 treatment reduced LV MLK3 protein expression (normalized to β-actin) compared with vehicle-treated littermates. After 14-day administration, URMC-099 induced significant increases in LV and heart mass, normalized to tibia length (TL), (LV/TL males: 4.21 ± 0.03 mg/mm vehicle vs 5.20 ± 0.10 mg/mm URMC-099; p&lt;0.01; females: 3.73 ± 0.14 mg/mm vehicle vs 4.30 ± 0.13 mg/mm URMC-099; p&lt;0.01). LV tissue from URMC-099-treated mice displayed a 7-fold increase in luciferase fluorescence (p&lt;0.001), compared with vehicle-treated, indicating increased NFAT activation by URMC-099. Conclusion: Systemic MLK3 inhibition with URMC-099 causes reduced MLK3 expression, LVH, and increased CM NFAT activity. Because URMC-099 does not raise blood pressure, our findings support that MLK3 functions as a tonic brake on NFAT activation in the CM and opposes cardiac hypertrophy through kinase-dependent mechanisms.
Abstract P2171: Lentivirus TAZ Gene Therapy Restores Metabolic Functions In Taz Ko Mice Model Of Barth Syndrome
Authors: Rahul Raghav, Junya Awata, Gregory Martin, Douglas Strathdee, Robert M Blanton, Michael T Chin
Journal: Circulation Research (2023)

Introduction: Barth Syndrome is an X-linked disorder caused by mutation in thegene encoding tafazzin (or TAZ ), characterized by dilated cardiomyopathy, muscularhypotonia and cyclic neutropenia. TAZ is a mitochondrial acyltransferase that generatesmature cardiolipin (CL) from monolysocardiolipin (MLCL) and is essential formitochondrial morphology and function. Patients with BTHS exhibit a characteristicreduction in the levels of CL and an accumulation of MLCL. Currently, there is no targetedtreatment exists for these patients. We characterized a global TAZ knockout mousemodel of BTHS and tested the efficacy of Lentivirus-mediated TAZ gene therapy. Methods: We generated third generation lentiviral constructs without an insert, withGreen Fluorescent Protein, with recombinant tafazzin and with recombinant tafazzinmodified to contain a cellular penetrating peptide (hTAZ-CPP) that promotes uptake intotafazzin-deficient cells. We tested the efficacy of these recombinant lentiviruses, first inmouse embryo fibroblasts (MEF) isolated from tafazzin knockout mice and then intafazzin knockout mice after systemic administration. Results: We have found that treatment with hTAZ-CPPs reduces the MLCL/CL ratio tonear wild type levels, thereby rescuing the defect in cardiolipin remodeling in TAZ KOMEFs. Serial echocardiograms revealed significant increase in septal wall thickness andsignificant decrease in ejection fraction, fractional shortening and global longitudinal strainin TAZ KO mice. The TAZ KO mice were treated systemic lentivirus at 5 weeks of ageand then echocardiograms were performed at every 4 weeks until 16 weeks. After thetreatment septal wall thickness continued to increase and ejection fraction, fractionalshortening and global longitudinal strain continued to decrease in the empty virus or GFPtreated mice, whereas they are remained stable in those treated with hTAZ-CPPs. Conclusion: These findings suggest that Lentivirus-mediated TAZ gene replacementtherapy enhanced with cell penetrating peptides has potential as a future treatment forBTHS.
Abstract P3033: Thrombospondin-1 Promotes Vascular Smooth Muscle Cell De-differentiation In Diabetes
Authors: Shreya Gupta, Amy Mathias, Jason Lallo, Neha Bhavnani, Priya Raman
Journal: Circulation Research (2023)

Diabetes is a major risk factor for atherosclerosis. Diabetic patients are prone to increased vascular smooth muscle cell (VSMC) migration and proliferation, hallmarks of VSMC de-differentiation, critical for the evolution of atherosclerosis. Multiple studies, including our earlier work, support the role of thrombospondin-1 (TSP1), a proatherogenic matricellular protein, as a putative link between diabetes and vascular disease. However, molecular mechanism(s) by which TSP1 promotes diabetic vasculopathy are incompletely understood. The goal of this work was to explore whether smooth muscle TSP1 (smTSP1) regulates VSMC phenotypic switch in diabetes. We generated conditional SMC-specific TSP1 knockout (smTSP1 KO ) mice by crossing female TSP1 fl/fl mice with tamoxifen (tmx)-inducible Myh11-CreER T2 male mice. For Cre recombination, 5-wks-old male TSP1 fl/fl Cre tg and age-matched TSP1 +/+ Cre tg littermate mice were treated with 60 mg/Kg/day tmx i.p. once daily for 5 consecutive days. To induce type 2 diabetes, mice cohorts were placed on western diet at 7-wks-age followed by streptozotocin (50 mg/Kg/day, i.p. x 5 days) at 16-wks-age. Mice were harvested at 20-wks-age for plasma and tissue collection. Immunoblotting of aortic lysates revealed significant decrease in TSP1 expression in tmx-treated TSP1 fl/fl Cre tg , validating smTSP1 KO mice. Lack of smTSP1 increased ACTA2 (SM contractile marker) expression by 2-fold accompanied with 50% reduction in CDK4 (cell cycle regulator) expression in diabetic smTSP1 KO vs. smTSP1 WT aortic vessels. Consistently, in murine aortic SMC (MASMC) primary cultures derived from smTSP1 KO mice, loss of smTSP1 augmented (&gt;65%) LMOD1 (SM contractile marker) and SRF (transcriptional activator of SM contractile genes) expression coupled with diminished (45%) vimentin (SM synthetic marker) expression in 25mM glucose-treated smTSP1 KO vs. smTSP1 WT cells. Further, propidium iodide staining via flow cytometry revealed reduced S-phase cell counts concomitant to increased G 0 -G 1 cell distribution in glucose-treated smTSP1 KO vs. smTSP1 WT MASMC. Together, our data demonstrates a direct role of TSP1 in VSMC phenotypic transition and opens novel avenues for cell-specific TSP1-based therapies for treatment of diabetes.